Mesenchymal stem or stromal cells (MSCs) are adult connective tissue progenitor cells with multilineage differentiation and anti-inflammatory properties. Due to these properties, MSCs are being evaluated as cellular therapy for regenerative medicine and inflammatory diseases. This proposal will address two critical issues related with MSCs biology; i) to understand the mechanisms of mesenchymal stem/stromal cell-mediated attenuation of acute lung injury in a relevant large animal model and ii) to examine if MSCs therapy influences the host immune response to influenza virus infection. Several studies in mouse models have suggested that MSCs attenuate lung injury and enhance lung repair. Most of the data of evaluation of MSCs as cellular therapy for human diseases have come from rodent models. However, the mechanisms of immunoregulation by mouse MSCs are different than those of human MSCs. Therefore, there is a need to conduct studies in clinically relevant animal model that closely resembles to humans in anatomy, physiology, body size and host response. In this application, we will use well defined LPS induced lung injury in pig as a model of human acute lung injury (ALI) to examine the mechanisms of MSC-mediated attenuation of ALI. First, we will isolate MSCs from bronchoalveolar lavage fluid (BAL) and bone marrow (BM). We will examine cell proliferation, colony formation, in vitro differentiation potential and their immunomodulatory properties. Next, we will examine mechanisms by which multipotent BAL/BM-MSCs attenuate acute lung injury and facilitate lung repair. We will exogenously administer in vitro expanded MSCs 24 hours after the onset of LPS-induced lung injury. We will examine the number of MSCs engrafting to the injured lung; effect on endogenous epithelial cell differentiation/proliferation; effect on macrophages, dendritic cells, T cells and NK cells functios and their effect on the attenuation of the disease and tissue repair and regeneration. In second objective, we will examine the effect of MSC-mediated immunoregulation in ALI in pig on anti-influenza immunity and virus clearance in pig model. Two, 4 and 6-weeks after MSCs therapy for ALI, we will infect pigs influenza virus. We will examine virus shedding, virus titers in lungs cytokines, chemokines, histological lung lesions, and virus- specific proliferating CD4 and CD8 T cells. The findings of these studies will provide new insights into the mechanisms of MSC-mediated beneficial effects in a relevant large animal model. Also, data obtained from this proposal will help to design preventive measures for the control of influenza virus infections in stem cell transplant patients.